Abstract
Four disulfide bridges of bovineα-lactalbumin (α-lact) were selectively reduced to obtain its derivatives with three, two, and zero disulfide bridges (designated as 3SS, 2SS, and OSSα-lact, respectively). The original helicity was almost maintained in 3SSα-lact missing only the Cys6-Cysl20 bridge. Upon the reduction of both Cys28-Cys111 and Cys6-Cys120 bridges, various changes occurred in the protein. In particular, the maximum fluorescence of 1-anilinonaphthalene-8-sulfonic acid was observed in this stage. Upon the reduction of all disulfide bridges, the hydrophobic box of the protein, formed by Trp60, Ile95, Tyr103, and Trp104, was disrupted and an internal helical structure was destroyed. The conformation of each derivative was examined mainly in a solution of sodium dodecyl sulfate. In the surfactant solution, the helicity increased from 33% to 37% in 3SSα-lact, from 26% to 31% in 2SSα-lact, and from 18% to 37% in OSSα-lact, as against from 34% to 44% in intactα-lact. On the other hand, the tryptophan fluorescence of each derivative was affected in very low surfactant concentrations, suggesting that the tertiary structure considerably changed prior to the secondary structural change in the surfactant solution.
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Acharya, K. R., Stuart, D. I., Walker, N. P. C., Lewis, M., and Phillips, D. C. (1989).J. Mol. Biol. 208, 99–127.
Acharya, K. R., Stuart, D. I., Phillips, D. C., and Scheraga, H. A. (1990).J. Protein Chem. 9, 549–563.
Acharya, K. R., Ren, J., Stuart, D. I., Phillips, D. C., and Fenna, R. E. (1991).J. Mol. Biol. 211, 571–581.
Browne, W. J., North, A. G T., Phillips, D. C., Brew, K., Vanaman, T. C., and Hill, R. L. (1969).J. Mol. Biol. 42, 65–86.
Chen, Y. H., Yang, J. T., and Chau, K. H. (1974).Biochemistry 13, 3350–3359.
Crestfleld, A. M., Moore, S., and Stein, W. H. (1963).J. Biol. Chem. 238, 622–627.
Dolgikh, D. A., Gilmanshin, R. I., Brazhnikov, E. V., Bychkova, V. E., Semisotnov, G. V., Venyaminov, S. Y., and Ptitsyn, O. B. (1981).FEBS Lett. 136, 311–315.
Ewbank, J. J., and Creighton, T. E. (1993a).Biochemistry 32, 3677–3693.
Ewbank, J. J., and Creighton, T. E. (1993b).Biochemistry 32, 3694–3707.
Hamada, S., and Takeda, K. (1993).J. Protein Chem. 12, 477–482.
Hiraoka, Y., Segawa, T., Kuwajima, K., Sugai, S., and Murai, N. (1980).Biochem. Biophys. Res. Commun. 95, 1098–1104.
Ikeguchi, M., and Sugai, S. (1989).Int. J. Peptide Protein Res. 33, 289–297.
Ikeguchi, M., Sugai, S., Fujino, M., Sugawara, T., and Kuwajima, K. (1992).Biochemistry 31, 12695–12700.
Iyer, K. S., and Klee, W. A. (1973).J. Biol. Chem. 248, 707–710.
Koga, K., and Berliner, L. J. (1985).Biochemistry 24, 7257–7262.
Kronman, M. J., and Andreotti, R. E. (1964).Biochemistry 3, 1145–1151.
Kuwajima, K., Nitta, K., Yoneyama, M., and Sugai, S. (1976).J. Mol. Biol. 106, 359–373.
Kuwajima, K., Hiraoka, Y., Ikeguchi, M., and Sugai, S. (1985).Biochemistry 24, 874–881.
Kuwajima, K., Ikeguchi, M., Sugawara, T., Hiraoka, Y., and Sugai, S. (1990).Biochemistry 29, 8240–8249.
Lala, A. K., and Kaul, P. (1992).J. Biol. Chem. 267, 19914–19918.
Nolting, B., Jiang, M., and Sugar, S. G. (1993).J. Am. Chem. Soc. 115, 9879–9882.
Schechter, Y., Patchornik, A., and Burstein, Y. (1973).Biochemistry 12, 3407–3413.
Takeda, K., and Moriyama, Y. (1990).J. Protein Chem. 9, 573–582.
Warme, P. K., Momany, F. A., Rumball, S. V., Tuttle, R. W., and Scheraga, H. A. (1974).Biochemistry 13, 768–782.
Weber, G., and Young, L. B. (1964).J. Biol. Chem. 239, 1415–1423.
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Takeda, K., Ogawa, K., Ohara, M. et al. Conformational changes of α-lactalbumin induced by the stepwise reduction of its disulfide bridges: The effect of the disulfide bridges on the structural stability of the protein in sodium dodecyl sulfate solution. J Protein Chem 14, 679–684 (1995). https://doi.org/10.1007/BF01886906
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DOI: https://doi.org/10.1007/BF01886906